Pituitary secretory vesicle enzymes involved in the processing of pro-opiomelano-cortin (POMC, pro-ACTH/endorphin) and pro- vasopressin were studied. A paired, basic residue specific prohormone converting enzyme (PCE) previously purified and characterized as an aspartyl protease was shown to be secreted from bovine intermediate lobe together with alpha-MSH, in a co- ordinately regulated manner. Pepstatin A, an inhibitor of PCE, blocked processing of POMC in the mouse intermediate lobe further supporting a physiological role of the enzyme in vivo. The POMC and provasopressin genes have been transfected into CV1 cells (green monkey kidney cell line) using a vaccinia virus transient expression system. The transfected cells secrete these prohormones in the intact form which can be recovered for use as substrate for PCE. An aminopeptidase B-like enzyme which removes basic residues from peptides cleaved by PCE was also shown to be co-secreted with alpha-MSH form bovine intermediate lobe cells in a regulated manner. The effect of salt- loading on POMC biosynthesis in the pituitary was studied. POMC mRNA levels, PIMC synthesis and plasma alpha-MSH levels decreased in the intermediate pituitary after 2 days of salt- loading and returned to normal after 9 days. In contrast, anterior pituitary POMC mRNA levels and plasma ACTH increased after 2 days of salt-loading and returned to normal by 4 days. In situ hybridization analyses of CRF and vasopressin (AVP) mRNA levels in hypothalamic neurons during salt loading showed no change in CRF but an increase in AVP mRNA levels. Studies on the interaction of AVP and CRF show that AVP is highly effective in attenuating CRF action at low doses of CRF.